莲腐败病菌pg基因的克隆及真核表达分析

Cloning and Eukaryotic Expression of pg1 Gene from Fusarium oxysporum f. sp. nelumbicola

针对尖孢镰刀菌莲专化型引起的莲腐败病是对白莲产业威胁最大的一种毁灭性病害。在植物病原菌侵染寄主植物引起病害的过程中多聚半乳糖醛酸酶(Polygalacturonase,PG)起着关键作用,是重要的致病因子。实验通过RT-PCR和RACE方法对莲腐败病菌的pg1全长cDNA进行克隆,获得一个编码371个氨基酸的完整开放阅读框。通过毕赤酵母表达系统成功构建了PG1同工酶的真核表达载体,利用甲醇诱导分泌出胞外蛋白PG1,分子量大小约为38 ku,与之前预测的分子量大小一致。研究结果为进一步分析PG基因的功能、探讨该基因在病原菌浸染过程中的作用机理和莲腐败病的防治提供了科学依据。

Rhizome rot of lotus,caused by Fusarium oxysporum Schl.f.sp.nelumbicola(Nis.&Wat.)Booth(FON),is a soil-borne disease,which has become the most destructive disease of lotus in recent years,and caused enormous economic loss.Polygalacturonase(PG)is one of the main enzymes that degrade the pectin of plant cell wall,playing an important role in the pathogenicity of plant pathogenic fungi.In this study,the full-length cDNA named pg1 of polygalacturonase gene was cloned by RT-PCR and RACE from Fusarium oxyporum f.sp.nelumbicola.And eukaryotic expression vector of pg1 was fabricated to research the enzymatic characteristics and pathogenic analysis of PG isozyme by Pichia pastoris expression system.The cloned pg1 consisted of a single open reading frame(ORF)encoding 371 amino acids.The protein molecular weight of PG1 was 38 ku by SDS-PAGE and Western blot detection.The results provided a foundation for studying the function of PG genes and understanding the molecular pathogenic mechanisms of FON.